Production of auto-anti-idiotypic antibody during the normal immune response to TNP-ficoll. II. Hapten-reversible inhibition of anti-TNP plaque-forming cells by immune serum as an assay for auto-anti-idiotypic antibody

Abstract

Sera taken from AKR/J mice 7 d [days] after the i.v. injection of 2,4,6-trinitrophenyl[TNP]-lys-Ficoll (TNP-F) caused a specific inhibition of anti-TNP plaque-forming cells (PFC) in vitro. This inhibition was reversed by the incorporation of 10-8-10-7 M TNP-.epsilon.-amino-n-caproic acid (TNP-EACA) into the agar during the PFC assay. The factor responsible for the hapten-reversible PFC inhibition was removed from serum by passage through an anti-immunoglobulin [Ig] column or through a 2,4,-dinitrophenyl-human-serum-albumin-bromoacetylcellulose [DNP-HSA-BAC] plus anti-TNP-antibody column, but not by DNP-HSA-BAC alone. This Ig-like substance, lacking anti-TNP activity but reacting with anti-TNP antibody of AKR/J origin, was most likely an auto-anti-idiotypic antibody that had been produced during the normal course of the response of AKR/J mice to TNP-F. Pools of anti-idiotypic-antibody-containing antisera inhibited anti-TNP plaque formation to varying degrees when tested on d-4 PFC from different mice of the same inbred strain, suggesting a variability in idiotype expression. Four days after transfer of immune (7 d after 10 .mu.g TNP-F, administered i.v.) AKR/J spleen cells plus 10 .mu.g TNP-F into syngeneic mice, the number of PFC detectable in the recipients'' spleens could be markedly augmented by the inclusion of TNP-EACA in the agar during the PFC assay. Incubation of spleen cells containing such hapten-augmentable PFC with TNP-EACA yielded a factor in the supernate that caused a specific, in vitro, hapten-reversible inhibition of anti-TNP PFC. Studies with immunoadsorbents indicated that this PFC-inhibiting factor was antigenically Ig-like, lacked anti-TNP-antibody activity, but reacted with anti-TNP antibody of AKR/J origin. The results are consistent with the view that this PFC inhibitor is auto-anti-idiotypic antibody that is involved in the normal regulation of the immune response. Hapten-reversible inhibition of plaque formation may be employed as an assay for anti-idiotypic antibody and the conditions for such an assay were described. The detection of hapten-augmentable PFC suggests the presence of auto-anti-idiotypic antibody.